Poster abstracts

Poster number 76 submitted by David Markwardt

A microarray screen for physiologic targets of the nonsense-mediated mRNA decay (NMD) pathway in Schizosaccharomyces pombe

David Markwardt (Department of Zoology, Ohio Wesleyan University)

Abstract:
Eukaryotes monitor mRNA transcript form and quality with different surveillance systems. One such system, called nonsense-mediated mRNA decay (NMD), can detect and help eliminate transcripts with premature termination codons (PTCs). The core components of the NMD pathway were first characterized in Saccharomyces cerevisiae and orthologs have since been identified in all other eukaryotes examined (Chang, 2007; Maquat, 2004). While the molecular mechanisms that underlie the selective degradation of PTC-containing transcripts are increasingly well understood, less is known about the source or nature of NMD substrates in wild-type cells. Various microarray and bioinformatics-based studies in S. cerevisiae, C. elegans, and mammalian cell culture suggest that PTC-containing products of alternative splicing may be an important class of physiologic substrate of NMD (Lewis, 2003; He, 2003; Wittmann, 2006). The recent identification and characterization of core NMD-factors in Schizosaccharomyces pombe have allowed us to undertake a systematic, microarray-based screen for S. pombe physiologic targets of NMD (Rodriguez-Gabriel, 2006). Using PCR-microarrays from the Stony Brook Spotted Microarray Facility, we have identified more than 100 transcripts at least two-fold more abundant in an NMD(-) background, compared to wild-type. The large number of intron-containing genes in the S. pombe genome (~43% compared to ~5% in S. cerevisiae) suggests that the pool of natural targets identified by our arrays might include alternatively spliced pre-mRNAs. We are currently using RT-PCR with primers flanking predicted splice junctions of each transcript in order to identify these putative NMD-targeted splice products. Studies such as this one will help us understand the role NMD plays in post-transcriptional regulation of gene expression.

References:
1. Chang YF, et al. Annu Rev Biochem. 2007; 76: 51-74
2. Maquat LE, Curr Genomics. 2004; 5: 175–190
3. Lewis BP, PNAS. 2003; 100(1): 189–192
4. He F, et al. Mol Cell. 2003; 12(6):1439-1452
5. Wittmann J, et al. Mol Cell Biol. 2006; 26(4): 1272-1287
6. Rodriguez-Gabriel MA, et al., MCB. 2006; 26(17): 6347–6356

Keywords: NMD, mRNA surveillance, yeast